Description: The fragile X syndrome is due to the absence of the FMR1 protein, FMRP. A mouse Finn knockout exhibits cognitive deficiencies and macroorchidism, recapitulating features of the human syndrome. Despite advances in our understanding of FMRP function and the consequence of its absence, many simple, yet fundamental questions remain elusive. A critical need throughout the entire gamut of such research is the availability of mice and cultured cells where Fmrp levels are easily modulated. The PL proposes here the development of several such models. Several lines of mice engineered where the Fmrp levels can be regulated by doxycycline exposure will be developed These will range from the traditional transgenic animal with the tetracycline-response element (TRE) promoter driving the Fmr1 cDNA on a knockout background to more sophisticated approaches. One novel approach maintains normal alternative splicing variation and normal cell expression specificity by replacing, in embryonic stem cells, the normal mouse FMR1 promoter with a TRE without otherwise modifying the murine gene. Another novel approach will be the fusion of a HIV-TAT transducing domain with the CRE recombinase. Purified fusion protein can easily enter all cell types and delete the genomic interval between direct lox repeats. Using this approach the PL will develop the ability to selectively ablate FMR1 in discreet organs and brain regions. From all mouse models developed, immortalized cell lines will also be developed for experiments where cultured cells would be most appropriate. All models will be rigorously tested for parameters of inducible FMR1 expression and decay. Using these models the PL will test fundamental issues involving fragile X syndrome, ranging from biochemical and cell biological questions to determining if FMRP expression is essential during development or if later expression of FMRP can influence the cognitive deficit. Such data is of fundamental importance in the consideration of therapeutic interventions. Developing, testing, and utilizing these mice and cells derived from them should substantially advance the analysis and understanding of FMRP function and the consequences of its absence.